Transcription factors c-Myc and CDX2 mediate E-selectin ligand expression in colon cancer cells undergoing EGF/bFGF-induced epithelial–mesenchymal transition

Sialyl Lewis x (sLex) and sialyl Lewis a (sLea) glycans are expressed on highly metastatic colon cancer cells. They promote extravasation of cancer cells and tumor angiogenesis via interacting with E-selectin on endothelial cells. Recently, epithelial–mesenchymal transition (EMT) has been noted as a critical phenotypic alteration in metastatic cancer cells. To address the association between sLex/a expression and EMT, we assessed whether sLex/a are highly expressed on colon cancer cells undergoing EMT. Treatment of HT29 and DLD-1 cells with EGF and/or basic FGF (bFGF) induced EMT and significantly increased sLex/a expression resulting in enhanced E-selectin binding activity. The transcript levels of the glycosyltransferase genes ST3GAL1/3/4 and FUT3 were significantly elevated and that of FUT2 was significantly suppressed by the treatment. We provide evidence that ST3GAL1/3/4 and FUT3 are transcriptionally up-regulated by c-Myc with probable involvement of Ser62 phosphorylation, and that FUT2 is transcriptionally down-regulated through the attenuation of CDX2. The contribution of c-Myc and CDX2 to the sLex/a induction was proved to be significant by knockdown or forced expression experiments. Interestingly, the cells undergoing EMT exhibited significantly increased VEGF secretion, which can promote tumor angiogenesis in cooperation with sLex/a. Finally, immunohistological study indicated high E-selectin ligand expression on cancer cells undergoing EMT in vivo, supporting their coexistence observed in vitro. These results suggest a significant link between sLex/a expression and EMT in colon cancer cells and a pivotal role of c-Myc and CDX2 in regulating sLex/a expression during EMT.

[1]  R. Kannagi,et al.  Contribution of carbohydrate antigens sialyl Lewis A and sialyl Lewis X to adhesion of human cancer cells to vascular endothelium. , 1993, Cancer research.

[2]  C. Bucana,et al.  Level and function of epidermal growth factor receptor predict the metastatic potential of human colon carcinoma cells. , 1995, Clinical cancer research : an official journal of the American Association for Cancer Research.

[3]  R. Cummings,et al.  Perspectives series: cell adhesion in vascular biology. Role of PSGL-1 binding to selectins in leukocyte recruitment. , 1997, The Journal of clinical investigation.

[4]  R. Cummings,et al.  Role of PSGL-1 binding to selectins in leukocyte recruitment. , 1997, The Journal of clinical investigation.

[5]  K Konstantopoulos,et al.  Perspectives Series: Cell Adhesion in Vascular Biology Effects of Fluid Dynamic Forces on Vascular Cell Adhesion , 1996 .

[6]  B. Zetter,et al.  Angiogenesis and tumor metastasis. , 1998, Annual review of medicine.

[7]  A. Jemal,et al.  Global cancer statistics , 2011, CA: a cancer journal for clinicians.

[8]  P. Carmeliet,et al.  Angiogenesis in cancer and other diseases , 2000, Nature.

[9]  Y Taya,et al.  Multiple Ras-dependent phosphorylation pathways regulate Myc protein stability. , 2000, Genes & development.

[10]  Hyung Jin Kim,et al.  Vascular Endothelial Growth Factor Expression of Intercellular Adhesion Molecule 1 (ICAM-1), Vascular Cell Adhesion Molecule 1 (VCAM-1), and E-selectin through Nuclear Factor-κB Activation in Endothelial Cells* , 2001, The Journal of Biological Chemistry.

[11]  Channing S. Mahatan,et al.  Expression of the gut-enriched Krüppel-like factor (Krüppel-like factor 4) gene in the human colon cancer cell line RKO is dependent on CDX2 , 2001, Oncogene.

[12]  N. Goldstein,et al.  Epidermal growth factor receptor immunohistochemical reactivity in patients with American Joint Committee on Cancer Stage IV colon adenocarcinoma , 2001, Cancer.

[13]  Y. Jeng,et al.  Tumor angiogenesis and its possible role in intravasation of colorectal epithelial cells. , 2001, Clinical cancer research : an official journal of the American Association for Cancer Research.

[14]  K. Kumamoto,et al.  Roles of cell adhesion molecules in tumor angiogenesis induced by cotransplantation of cancer and endothelial cells to nude rats. , 2002, Cancer research.

[15]  Hiroshi Yamamoto,et al.  Homeodomain protein CDX2 regulates goblet-specific MUC2 gene expression. , 2003, Biochemical and biophysical research communications.

[16]  K. Kumamoto,et al.  Hypoxia induces adhesion molecules on cancer cells: A missing link between Warburg effect and induction of selectin-ligand carbohydrates. , 2004, Proceedings of the National Academy of Sciences of the United States of America.

[17]  R. Kannagi,et al.  Carbohydrate‐mediated cell adhesion in cancer metastasis and angiogenesis , 2004, Cancer science.

[18]  K. Kumamoto,et al.  Loss of Disialyl Lewisa, the Ligand for Lymphocyte Inhibitory Receptor Sialic Acid-Binding Immunoglobulin-Like Lectin-7 (Siglec-7) Associated with Increased Sialyl Lewisa Expression on Human Colon Cancers , 2004, Cancer Research.

[19]  R. Kannagi.  Carbohydrate-mediated cell adhesion involved in hematogenous metastasis of cancer , 1997, Glycoconjugate Journal.

[20]  S. Eccles,et al.  Plasma basic fibroblast growth factor levels in colorectal cancer: A clinically useful assay? , 2004, Clinical & Experimental Metastasis.

[21]  E. Suh,et al.  The role of Cdx proteins in intestinal development and cancer , 2004, Cancer biology & therapy.

[22]  M. Loda,et al.  c-Myc phosphorylation is required for cellular response to oxidative stress. , 2006, Molecular cell.

[23]  Boris Pasche,et al.  TGF-β signaling alterations and susceptibility to colorectal cancer , 2007 .

[24]  Boris Pasche,et al.  TGF-beta signaling alterations and susceptibility to colorectal cancer. , 2007, Human molecular genetics.

[25]  T. Brabletz,et al.  The intestine-specific homeobox gene Cdx2 decreases mobility and antagonizes dissemination of colon cancer cells , 2008, Oncogene.

[26]  D. Radisky,et al.  Epithelial-mesenchymal transition and the stem cell phenotype. , 2008, Cell stem cell.

[27]  Wenjun Guo,et al.  The Epithelial-Mesenchymal Transition Generates Cells with Properties of Stem Cells , 2008, Cell.

[28]  Sabine Tejpar,et al.  Implications for KRAS status and EGFR-targeted therapies in metastatic CRC , 2009, Nature Reviews Clinical Oncology.

[29]  R. Huang,et al.  Epithelial-Mesenchymal Transitions in Development and Disease , 2009, Cell.

[30]  M. Henriksson,et al.  Phosphorylation by Cdk2 is required for Myc to repress Ras-induced senescence in cotransformation , 2009, Proceedings of the National Academy of Sciences.

[31]  Y. Nagano,et al.  Overexpression of the fibroblast growth factor receptor-1 gene correlates with liver metastasis in colorectal cancer. , 2009, Oncology reports.

[32]  Ying Liu,et al.  Gastrointestinal differentiation marker Cytokeratin 20 is regulated by homeobox gene CDX1 , 2009, Proceedings of the National Academy of Sciences.

[33]  K. Handa,et al.  Specific glycosphingolipids mediate epithelial-to-mesenchymal transition of human and mouse epithelial cell lines , 2009, Proceedings of the National Academy of Sciences.

[34]  Raghu Kalluri,et al.  The basics of epithelial-mesenchymal transition. , 2009, The Journal of clinical investigation.

[35]  Shuji Ogino,et al.  Relationship of CDX2 Loss with Molecular Features and Prognosis in Colorectal Cancer , 2009, Clinical Cancer Research.

[36]  M. Aoki,et al.  CDX transcription factors positively regulate expression of solute carrier family 5, member 8 in the colonic epithelium. , 2010, Gastroenterology.

[37]  Kohei Miyazono,et al.  TGFβ signalling: a complex web in cancer progression , 2010, Nature Reviews Cancer.

[38]  R. Kannagi,et al.  Epigenetic silencing of the sulfate transporter gene DTDST induces sialyl Lewisx expression and accelerates proliferation of colon cancer cells. , 2010, Cancer research.

[39]  K. Handa,et al.  Functional role of gangliotetraosylceramide in epithelial‐to‐mesenchymal transition process induced by hypoxia and by TGF‐β , 2010 .